Stressors affect the use of a wide range of substances involved in drug abuse; for example, stressful environments have been reported to stimulate cigarette smoking. Stressors may affect drug self-administration (SA) through their diverse effects on stress-responsive CNS systems, such as those regulating neurohormonal and noradrenergic function within the hypothalamus (e.g. paraventricular nucleus; PVN) and brainstem, respectively. We hypothesize that chronic nicotine SA, acting as a stressor, activates and modifies these stress-responsive systems, leading to changes in their stress-responsiveness. The specific aims of this application are focused on characterizing and elucidating mechanisms underlying these changes in HPA and PVN stress-responsivity induced by chronic nicotine SA. A unique model of daily (23 h/d) unlimited access to nicotine SA will be used to study the effects of nicotine on HPA and PVN responses to stressors. Spec. Aim 1 will characterize the effects of chronic nicotine SA on hypothalamo-pituitary-adrenal (HPA) activation (plasma ACTH/corticosterone) and PVN norepinephrine release by a range of stressors at different intensities. In Spec. Aim 2, mechanisms responsible for the altered stress-responsiveness of chronic nicotine SA rats will be determined. These will include studies on the effects of chronic nicotine SA on PVN neuropeptide mRNA expression, the activation of PVN c-fos expression by stressors, and the role of PVN noradrenergic inputs in the PVN and HPA responses to stressors. Complementary studies will also identify nicotinic cholinergic receptors that mediate activation of the brainstem-PVN noradrenergic axis by nicotine. Spec. Aim 3 will characterize the role of glutamate and NTS NMDA receptors in (1) nicotine-induced PVN NE secretion and HPA activation and in (2) the reduction of stress-induced PVN NE release caused by chronic nicotine SA. Preliminary observations indicate that specific NMDA receptor subunits in the nucleus tractus solitarius (NTS), with noradrenergic projections to the PVN, are downregulated by nicotine SA and that brainstem glutamate is necessary for the PVN NE response to nicotine. Therefore, studies in this section will involve the quantitation of specific NTS NMDA receptor subunits by immunoblotting. The functional effects of NMDA receptor downregulation also will be characterized by measuring PVN NE responses and the induction of NTS cfos expression by NMDA during nicotine SA. We postulate that NTS glutamate may be involved in stress-induced PVN NE secretion. Therefore, studies will evaluate whether inhibition of NTS NMDA receptors attenuates the PVN NE response to an acute stressor. Together, the studies in this proposal will clarify the effects of chronic nicotine SA on HPA responsiveness to stressors and on underlying adaptive changes in the brainstem HPA axis.